Use the links and their instructions from the Applications section to help answer
the following questions. Many
applications may need to be used at one time. Hint: Keep all results from the applications open
until problem set is complete, you will most likely need them more than once.
1. Find the most probable origin of the following nucleotide sequence. What is the degree of similarity (percent identity) between the top five vertebrae alignments and the nucleotide sequence?
Use WU-blastn or NCBI’s blastn (Sequence Identification Application).
MOST PROBABLE ORIGIN: Homo sapiens solute carrier family 5 (sodium/glucose cotransporter), member 1 (SLC5A1), mRNA. MIM# 182380
TOP FIVE VERTEBRAE ALIGNMENTS:
1. Homo sapien (human) ~ 98% identity
2. Pig ~ 74% identity,
3. Ovis aries(sheep) ~ 88% identity
4. Rattus novaegicus(rat) ~ 84% identity
5. Mus musculus(mouse) ~ 83% identity
OTHERS (depending on search): bos tarus(cow) ~ 89% identity, gallus gallus (chicken) ~ 75% identity, oryctolagus cuniculus(rabbit) ~ 80% identity
NON VERTEBRAE:drosophila(fruit fly), xylella fastidosa(bacteria)
2. What reading frame is this sequence read?
Use Molecular Biology Shortcuts (Translation Application) and find the reading frame (using 6 phases). Sometimes can also be seen in results from blastn (Sequence Identification).
READING FRAME: 2nd reading frame
3. What is the function of the protein product from this DNA?
See the blastn results.
PROTEIN FUNCTION: sodium/glucose cotransporter, used mostly in the brush border of the intestinal epithelium.
4. Do the restriction enzymes MaeIII, EcorI, MamI, and RcaI cut this sequence, and if so how many times do they cut, where do they cut (give approx. base pair region), and what is their recognition sequence? How does it compare with the original Human gene?
Use REBASE restriction mapping tool (Restriction Enzyme
Mapping in Applications).
|
Enzyme
Name |
No. cuts |
Positions
of cuts |
Recognition
sequence |
|
MaeIII |
5 |
41 237 1364
1963 2249 |
/gtnac |
|
MamI |
3 |
846 1286
1653 |
gatnn/nnatc |
|
RcaI |
2 |
1149
1248 |
t/catga |
|
EcoRI |
1 |
384 |
g/aattc |
HUMAN DNA (Homo sapiens solute carrier family 5 (sodium/glucose
cotransporter)):
|
Enzyme
Name |
No. cuts |
Positions
of cuts |
Recognition
sequence |
|
MaeIII |
6 |
46 239 385
1391 1966 2246 |
/gtnac |
|
MamI |
3 |
860 1313
1671 |
gatnn/nnatc |
|
RcaI |
2 |
1179 1275 |
t/catga |
|
EcoRI |
0 |
did not cut |
g/aattc |
5. To what human chromosome does this gene map?
Use OMNI in
Literature
CHROMOSOME: 22q13.1
6. What human dysfunction is related to this protein product? What are symptoms of this disease and is there a known treatment? List three full citations for articles that tell about the disorder related to this gene.
Use OMIM (Literature Searches) and enter either the protein family (SLC5A1) or the MIM# (182380) that you retrieved from the blastn. When you search for these, you should get back a list of related articles and their abstracts.
DYSFUNCTION: Glucose Galactose malabsorption.
SYMPTOMS: first seen in first weeks of life. Observe severe diarrhea and dehydration (usually fatal)
TREATMENT: eliminate glucose and galactose from the diet.
JOURNAL
ARTICLES: see OMNI on NCBI
7. Find a similar structure for this protein (You may not be able to find vertebrae structure). Give the name of the protein structure and the organism from which it comes.
Use Protein Data Base (Instructions on Obtaining Structure
files).
PROTEIN/ORGANISM: Glucose Permease/ Escherichia Coli
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